1. Field of the Invention
This invention relates to phenylcycloalkylmethylamino and phenylalkenylamino derivatives, including 1-phenyl-2-aminomethylcyclopropanes, that are modulators of melanin concentrating hormone type 1 (MCH 1) receptors. This invention also relates to pharmaceutical compositions comprising such compounds.
2. Description of the Related Art
Melanin concentrating hormone, or MCH, is a cyclic 19 amino acid neuropeptide that is produced within the hypothalamus of many vertebrate species including man. I.C.V. injection of MCH into the lateral ventricle of the hypothalamus has been shown to increase caloric consumption in rats over similarly treated control animals. Furthermore, rats having the ob/ob genotype exhibit a 50-80% increase in MCH mRNA expression as compared to leaner ob/+ genotype mice. MCH knockout mice are leaner than their MCH-producing siblings due to hypophagia and an increased metabolic rate. Thus, MCH is thought to be an important regulator of feeding behavior and body weight.
The MCH 1 receptor was originally obtained from human cDNA and genomic libraries and characterized as a 402 amino acid G-coupled protein receptor having substantial sequence identity to the somatostatin receptors. This receptor was named the SLC-1 receptor. A rat orthologue of the MCH 1 receptor was isolated from a rat brain cDNA library by Lakaye, et al. (BBA (1998) 1401: 216-220) and found to encode a 353 amino acid protein having seven transmembrane alpha helices and three consensus N-glycosylation sites. The rat MCH 1 receptor reported by Lakaye also disclosed was homologous to the human MCH 1 receptor disclosed earlier except for the removal of a 5xe2x80x2 intron. Accordingly, Lakaye, et al., deduced the xe2x80x9ccorrectedxe2x80x9d amino acid sequence of the N-terminus of MCH 1 receptor is found within a sequence deposited for a 128 kb fragment of human chromosome 22 encompassing the earlier disclosed MCH 1 receptor gene (Genbank accession number: Z86090).
The earlier reported 402 amino acid MCH 1 receptor protein does not interact with MCH. Thus, the 353 amino acid receptor first reported by Lakaye, is now considered to be the correct full-length sequence for the human MCH 1 receptor.
Immunohistochemistry studies of rat brain sections indicate that the MCH 1receptor is widely expressed in the brain. MCH 1 receptor expression was found in the olfactory tubercle, cerebral cortex, substantia nigra, basal forebrain CA1, CA2, and CA3 field of the hippocampus, amygdala, and in nuclei in the hypothalamus, thalamus, midbrain and hindbrain. Strong signals have been observed in the ventromedial and dorsomedial nuclei of the hypothalamus, two areas of the brain known to be involved in feeding behavior.
Upon binding MCH, MCH 1 receptors expressed in HEK 293 cell mediate a dose dependent release of intracellular calcium. Cells expressing MCH receptors have also been shown to exhibit a pertussis toxin sensitive dose-dependent inhibition of forskolin-elevated cyclic AMP, indicating that the receptor couples to a Gi/o G-protein alpha subunit.
Because MCH has been shown to be an important regulator of food intake and energy balance, ligands capable of modulating the activity of the MCH 1 receptor are highly desirable for the treatment of eating disorders and metabolic disorders. Orally available, small molecule, non-peptide antagonists of the MCH 1 receptor are particularly sought for the treatment of obesity.
The invention provides novel compounds, particularly phenylcycloalkylmethylamino and phenylalkenylamino compounds, including 1-phenyl-2-aminomethylcyclopropanes, that are small molecule MCH receptor ligands, especially MCH 1 receptor ligands, that are non-peptide and amino acid free, which compounds exhibit a Ki at the MCH receptor of less than 1 micromolar. Preferred MCH 1 receptors are mammalian receptors, including human and monkey MCH receptors and may either be cloned, recombinantly expressed receptors or naturally expressed receptors.
In certain embodiments these compounds also possess one or more, and preferably two or more, three or more, or all of the following properties in that they are: 1) multi-aryl in structure (having a plurality of un-fused or fused aryl groups), 2) orally available in vivo (such that a sub-lethal or pharmaceutically acceptable oral dose can provide a detectable in vivo effect such as a reduction of appetite), 3) capable of inhibiting the binding of MCH to the MCH receptor at nanonmolar concentrations or 4) capable of inhibiting the binding of MCH to the MCH receptor at sub-nanomolar concentrations.
The invention also provides novel compounds of Formula I, shown below, that bind specifically, and preferably with high affinity, to MCH receptors.
The invention also provides pharmaceutical compositions comprising compounds of Formula I together with at least one pharmaceutically acceptable carrier. The compounds are particularly useful in the treatment of metabolic, feeding, and sexual disorders. The invention further comprises a method of treating a patient in need of such treatment with a sufficient concentration of a compound of the invention. A preferred concentration is one sufficient to inhibit the binding of MCH to MCH 1 receptors in vitro. Treatment of humans, domesticated companion animals (pets) or livestock animals suffering such conditions with an effective amount of a compound of the invention is contemplated by the invention.
Also included in the invention are methods of treating eating disorders, particularly obesity and bulimia nervosa, comprising administering to a patient in need of such treatment a MCH 1 receptor modulator together with leptin, a leptin receptor agonist, or a melanocortin receptor 4 (MC4) agonist.
In a separate aspect, the invention provides methods of using compounds of this invention as positive controls in assays for receptor activity and using appropriately labeled compounds of the invention as probes for the localization of receptors, particularly MCH receptors, in tissue sections.
The invention provides compounds and compositions that are useful as inhibitors of MCH binding to MCH 1 receptor, and as inhibitors of MCH mediated signal transduction (e.g., they may be used as standards in assays of MCH binding and MCH-mediated signal transduction). The invention additionally comprises methods of inhibiting MCH binding to MCH receptors in vivo, preferably MCH 1 receptors present in the hypothalamus.
Accordingly, a broad embodiment of the invention is directed to a compounds and pharmaceutically acceptable salts of Formula I: 
wherein:
Q is a group of the Formula: 
xe2x80x83wherein
A is C1-C5 alkylene optionally mono-, di, or trisubstituted with substitutuents independently chosen from C1-C3 alkyl, C1-C3 alkoxy, halogen, halo(C1-C3)alkyl, halo(C1-C3)alkoxy, hydroxy, amino, and mono- or di(C1-C3)alkylamino;
R1, R2, R3, R4, R5, R6, R7, and R8 are the same or different and represent hydrogen, halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkyl amino, halo(C1-C6)alkyl, halo(C1-C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94SO2NH2, mono or dialkylsulfonamido, xe2x80x94C(O)NH2, or mono or di(C1-C6)alkylcarboxamido;
R9, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 independently represent hydrogen or C1 -C6 alkyl;
W is nitrogen or Cxe2x80x94Ra where Ra represents hydrogen, hydroxy, C1-C6 alkoxy, C1-C6 alkyl or cyano;
X represents halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkylamino, halo(C1-C6)alkyl, halo(C1-C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94CONH2, mono- or di(C1-C6)alkylcarboxamido, xe2x80x94SO2NH2, mono or di(C1-C6)alkylsulfonamido; or
X represents phenyl which may be optionally substituted by up to five substituents, which may be the same or different and are selected from the group consisting of hydrogen, halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkyl amino, halo(C1-C6)alkyl, halo(C1 -C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94CONH2, mono- or di-(C1 -C6)alkylcarboxamido, xe2x80x94SO2NH2, and mono or di(C1-C6)alkylsulfonamido;
Y is oxygen, sulfur, xe2x80x94S(O)xe2x80x94, or xe2x80x94SO2xe2x80x94; and
Z is C1-C6 alkyl or mono, di or trifluoromethyl.
The invention also provides intermediates and methods useful for preparing the compounds of Formula I.
The invention particularly includes compounds and salts of Formula I wherein Q is a ring and A is methylene optionally substituted with C1-C2 alkyl.
The invention is also specifically directed to compounds and salts of Formula I wherein W is nitrogen or CH and A is methylene. Preferred compounds and salts of this class are those wherein R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen. Other preferred compounds and salts of this class are those wherein R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen, X is halogen; Y is oxygen; and Z is C1-C6 alkyl. Also preferred are compounds and salts of Formula I wherein W is nitrogen or CH and A is methylene, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen, R1, R2, R3, R4, R5, R6, R7, and R8 may be the same or different and represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy; X is hydrogen, halogen, or phenyl, or most preferably X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
Particularly provided by the invention are compounds of Formula II 
and the pharmaceutically acceptable salts thereof; wherein
A is methylene optionally substituted with C1-C2 alkyl, and R1-R19, W, X, Y, and Z are as defined for Formula I.
Preferred compounds and salts of Formula II are those wherein W is nitrogen or CH.
Other preferred compounds and salts of Formula II are those wherein W is nitrogen or CH and R10, R11, R12, R13, R15, R17, R18, and R19 are hydrogen.
Also preferred are compounds and salts of Formula II wherein W is nitrogen or CH, R10, R11, R12, R13, R15, R17, R18, and R19 are hydrogen, R1, R2, R3, R4, R5, R6, R7, and R8 independently represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy; R14 and R16 are the same or different and are either hydrogen or methyl; X is hydrogen, halogen, or phenyl; Y is oxygen; and Z is C1-C6 alkyl.
Particularly preferred compounds and salts of Formula II are those wherein W is nitrogen or CH, R1, R6, R7, R8, R9, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen,
R2, R3, R4, and R5 are independently hydrogen, C1-C2 alkyl, C1-C2 alkoxy, or halogen; X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
The invention further provides compounds of Formula III 
and the pharmaceutically acceptable salts thereof, wherein R1-R19, W, X, Y, and Z are as defined for Formula I.
Preferred compounds and salts of Formula III are those wherein R13, R15, R17, R19, are hydrogen; and R10, R11, R12, R14, R16, and R18 independently represent hydrogen or methyl, or more preferably hydrogen.
Also preferred are compounds and salts of Formula III, wherein R10-R19 are hydrogen, and W is N or CH.
More preferred compounds and salts of Formula III are those wherein R10-R19 are hydrogen, W is N or CH; R1, R2, R3, R4, R5, R6, R7, and R8 independently represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy; X is hydrogen or halogen; Y is oxygen; and Z is C1-C6 alkyl.
Particularly preferred compounds and salts of Formula III are those wherein R10-R19 are hydrogen, W is N or CH, R1, R2, R3, R4 independently represent hydrogen, halogen, C1-C2alkyl, or C1-C2 alkoxy; R5, R6, R7, and R8 are hydrogen; X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
Another embodiment of the invention is directed to compounds and salts of Formula IV 
and the pharmaceutically acceptable salts thereof, wherein R1-R19, W, X, Y, and Z are as defined for Formula I.
Preferred compounds and salts of Formula IV are those wherein R13, R15, R17, R19, are hydrogen; and R10, R11, R12, R14, R16, and R18 independently represent hydrogen or methyl, or more preferably hydrogen.
Also preferred are compounds and salts of Formula IV, wherein R10-R19 are hydrogen, and W is N or CH.
More preferred compounds and salts of Formula IV are those wherein R10-R19 are hydrogen, W is N or CH; R1, R2, R3, R4, R5, R6, R7, and R8 independently represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy; X is hydrogen or halogen; Y is oxygen; and Z is C1-C6 alkyl.
Particularly preferred compounds and salts of Formula IV are those wherein R10-R19 are hydrogen, W is N or CH, R1, R2, R3, R4 independently represent hydrogen, halogen, C1-C2alkyl, or C1-C2 alkoxy; R5, R6, R7, and R8 are hydrogen; X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
The invention also provides compounds of Formula V 
or a pharmaceutically acceptable salt thereof wherein:
Q is a group of the Formula: 
xe2x80x83wherein:
A is C1-C5 alkylene optionally mono-, di, or trisubstituted with substitutuents independently chosen from C1-C3 alkyl, C1-C3 alkoxy, halogen, halo(C1-C3)alkyl, halo(C1-C3)alkoxy, hydroxy, amino, and mono- or di(C1-C3)alkylamino;
R1, R2, R3, R4, R5, R6, R7, and R8 are the same or different and represent hydrogen, halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkyl amino, halo(C1-C6)alkyl, halo(C1-C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94SO2NH2, mono or dialkylsulfonamido, xe2x80x94C(O)NH2, or mono or di(C1-C6)alkylcarboxamido;
R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 independently represent hydrogen or C1-C6 alkyl;
W is nitrogen or Cxe2x80x94Ra where Ra represents hydrogen, hydroxy, C1-C6 alkoxy, C1-C6 alkyl or cyano;
X represents halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkylamino, halo(C1-C6)alkyl, halo(C1-C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94CONH2, mono- or di(C1-C6)alkylcarboxamido, xe2x80x94SO2NH2, mono or di(C1-C6)alkylsulfonamido; or
X represents phenyl which may be optionally substituted by up to five substituents, which may be the same or different and are selected from the group consisting of hydrogen, halogen, cyano, nitro, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C1-C6 alkylthio, hydroxy, amino, mono or di(C1-C6)alkyl amino, halo(C1-C6)alkyl, halo(C1-C6)alkoxy, C1-C6 alkanoyl, C1-C6 alkoxycarbonyl, xe2x80x94COOH, xe2x80x94CONH2, mono- or di-(C1-C6)alkylcarboxamido, xe2x80x94SO2NH2, and mono or di(C1-C6)alkylsulfonamnido;
Y is oxygen, sulfur, xe2x80x94S(O)xe2x80x94, or xe2x80x94SO2xe2x80x94; and
Z is C1-C6 alkyl or mono, di or trifluoromethyl.
Compounds of Formula V are intermediates, useful in preparing compounds MCH 1 receptor ligands.
Preferred compounds of Formula V are those wherein Q is a group the formula 
where A is methylene optionally substituted with C1-C2 alkyl or A is a single bond. Such compounds will be referred to as compounds of Formula VA.
The invention is particularly directed to compounds of Formula VA wherein W is nitrogen or CH.
More preferred compounds of Formula VA are those wherein W is nitrogen or CH, and R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen.
Other preferred compounds of Formula VA are those wherein W is nitrogen or CH, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen, X is halogen; Y is oxygen and Z is C1-C6 alkyl.
Especially preferred compounds of Formula VA are those wherein W is nitrogen or CH, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen, R1, R2, R3, R4, R5, R6, R7, and R8 may be the same or different and represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy; X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
Particularly preferred compounds of Formula V include those where Q is a group of the formula: 
where A is methylene optionally substituted with C1-C2 alkyl. These compounds are hereinafter referred to as compounds of Formula VI-A. Specific compounds of Formula VI-A include those where A is methylene and R10 and R11 are methyl or, preferably, hydrogen.
Specific compounds of VA include those wherein W is nitrogen or CH. Preferred compounds of V and VA include those wherein R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are hydrogen.
Other specific compounds of VA include those wherein: X is halogen; Y is oxygen; and Z is C1-C6 alkyl.
Still other specific compounds of VA include those where
R1, R2, R3, R4, R5, R6, R7, and R8 are the same or different and represent hydrogen, halogen, C1-C6 alkyl, C1-C6 alkoxy, trifluoromethyl, or trifluoromethoxy;
X is halogen;
Y is oxygen; and
Z is C1-C6 alkyl.
Preferably not more than 5, and more preferably not more than 3, cyano or nitro groups are present in compounds of Formula I-Formula VA. Preferably not more than 2 of R1, R2, R3, R4, R5 are non-hydrogen substituents. Preferably not more than 5 and more preferably not more than 3 of R6, R7, R8, R9, R10, R11, R12, R13, R14, R15, R16, R17, R18, and R19 are non-hydrogen substituents.
Representative compounds of Formula I are shown in Table 1.
In certain situations, the compounds of this invention may contain one or more asymmetric carbon atoms, so that the compounds can exist in different stereoisomeric forms. These compounds can be, for example, racemates or optically active forms. In these situations, the single enantiomers, i.e., optically active forms, can be obtained by asymmetric synthesis or by resolution of the racemates. Asymmetric synthesis of compounds of the invention may be performed using the methods illustrated in Example 1, below. For compounds having an alpha-methyl benzyl group (R3 is methyl, R4 is hydrogen) the R enantiomer is preferred. Resolution of the racemates can be accomplished, for example, by conventional methods such as crystallization in the presence of a resolving agent, or chromatography, using, for example a chiral HPLC column.
Representative compounds of the invention, which are encompassed by Formula I, include, but are not limited to the compounds in Table I and their pharmaceutically acceptable acid addition salts. In addition, if the compound of the invention is obtained as an acid addition salt, the free base can be obtained by basifying a solution of the acid salt. Conversely, if the product is a free base, an addition salt, particularly a pharmaceutically acceptable addition salt, may be produced by dissolving the free base in a suitable organic solvent and treating the solution with an acid, in accordance with conventional procedures for preparing acid addition salts from base compounds.
Non-toxic pharmaceutical salts include salts of acids such as hydrochloric, phosphoric, hydrobromic, sulfuric, sulfinic, formic, toluenesulfonic, methanesulfonic, nitric, benzoic, citric, tartaric, maleic, hydroiodic, alkanoic such as acetic, HOOCxe2x80x94(CH2)nxe2x80x94COOH where n is 0-4, and the like. Those skilled in the art will recognize a wide variety of non-toxic pharmaceutically acceptable addition salts.
The invention also encompasses the acylated prodrugs of the compounds of Formula I. Those skilled in the art will recognize various synthetic methodologies which may be employed to prepare non-toxic pharmaceutically acceptable addition salts and acylated prodrugs of the compounds encompassed by Formula I.
Where a compound exists in various tautomeric forms, the invention is not limited to any one of the specific tautormers. The invention includes all tautomeric fonns of a compound.
This invention relates to compounds that bind with high affinity to the melanin concentrating hormone receptors, including human melanin concentrating hormone receptors. This invention also includes such compounds that bind with high selectivity to the melanin concentrating hormone receptors, including human and monkey melanin concentrating hormone receptors. Without wishing to be bound to any particular theory, it is believed that the interaction of the compounds of Formula I with the melanin concentrating hormone receptor results in the pharmaceutical utility of these compounds.
The invention further comprises methods of treating patients in need of such treatment with an amount of a compound of the invention sufficient to alter the symptoms of a disorder.
The diseases and/or disorders that can also be treated using compounds and compositions according to the invention include, but are not limited to, eating disorders, sexual disorders, obesity, bulimia, anorexia, diabetes, heart disease, stroke, anorgasmia, or psychogenic impotence.
The invention also provides pharmaceutical compositions comprising at least one compound of the invention together with at least one pharmaceutically acceptable carrier or excipient. Such pharmaceutical compositions include packaged pharmaceutical compositions for treating disorders responsive to melanin concentrating hormone receptor modulation, e.g., treatment of eating disorders such as obesity or bulimia or treatment of sexual disorders such as anorgasmic or psychogenic impotence. The packaged pharmaceutical compositions include a container holding a therapeutically effective amount of at least one melanin concentrating hormone receptor modulator as described supra and instructions (e.g., labeling) indicating that the contained composition is to be used for treating a disorder responsive to melanin concentrating hormone receptor modulation in the patient.
The invention also pertains to methods of inhibiting the binding of melanin concentrating hormone to melanin concentrating hormone receptors which methods involve contacting a compound of the invention with cells expressing melanin concentrating hormone receptors, wherein the compound is present at a concentration sufficient to inhibit melanin concentrating hormone binding to melanin concentrating hormone receptors in vitro. This method includes inhibiting the binding of melanin concentrating hormone to melanin concentrating hormone receptors in vivo, e.g., in a patient given an amount of a compound of Formula I that would be sufficient to inhibit the binding of melanin concentrating hormone to melanin concentrating hormone receptors in vitro. The amount of a compound that would be sufficient to inhibit the binding of melanin concentrating hormone to the melanin concentrating hormone receptor in vitro may be readily determined via a melanin concentrating hormone receptor binding assay, such as the assay described in Example 5. The membranes, comprising melanin concentrating hormone receptors, used to determine in vitro binding may be obtained from a variety of sources, for example from preparations of HEK 293 cells expressing cloned human or cloned monkey melanin concentrating hormone receptors, especially HEK 293 cells expressing such receptors.
The invention also pertains to methods for altering the signal-transducing activity of MCH receptors, particularly the MCH receptor-mediated release of intracellular calcium, said method comprising exposing cells expressing such receptors to an effective amount of a compound of the invention. This method includes altering the signal-transducing activity of MCH receptors in vivo, e.g., in a patient given an amount of a compound of Formula I that would be sufficient to alter the signal-transducing activity of MCH receptors in vitro. The amount of a compound that would be sufficient to alter the signal-transducing activity of MCH receptors may be determined via a MCH receptor signal transduction assay, such as the calcium mobilization assay described in Example 6.
The melanin concentrating hormone receptor ligands provided by this invention and labeled derivatives thereof are also useful as standards and reagents in determining the ability of a potential pharmaceutical to bind to the melanin concentrating hormone receptor.
Labeled derivatives the melanin concentrating hormone receptor ligands provided by this invention are also useful as radiotracers for positron emission tomography (PET) imaging or for single photon emission computerized tomography (SPECT).
Preferred compounds of the invention do not exhibit fungicidal activity. Such a lack of fungicidal activity may be demonstrated by no more than a 40% reduction of colony size (when treated with the compound at 100 p.p.m. and compared to untreated controls) of Aspergillus nidulans strain R153 when grown for 48 hours at 32xc2x0 C. on solid MAG medium. Optionally, BENOMYL, 100 p.p.m., may be used as a positive control. MAG medium is 2% malt extract, 0.2% peptone, 1% glucose and trace elements, pH 6.5. Trace elements as a 5000-fold concentrate consist of 10 g/l EDTA, 4.4 g/l ZnSO4.7H2O, 1.01 g/l MnCl2.4H2 O, 0.32 g/l CoCl2.6H2O, 0.315 g/l CuSO4.5H2O, 0.22 g/l (NH4)6Mo7O24.H2O, 1.47 g/l CaCl2.2H2O and 1.0 g/l FeSO4.7H2O. Medium is made solid by the addition of 1.5% agar.
Alternatively, such a lack of fungicidal activity may be demonstrated by an infection frequency of 60-100% (as compared to untreated plants) for each of Puccinia recondita (leaf rust) on wheat, Erysiphe graininis (powdery mildew) on barley, Venturia inaequalis (scab, black spot) on apple plants, and Cercospora arachidicola (early leafspot) on peanut.
The technique employed to determine fungicidal activity is as follows. The plants are grown in John Innes Potting Compost (No. 1, or Seed, as appropriate) in 4 cm diameter mini-pots. A layer of fine sand is placed at the bottom of the pot to facilitate uptake of test compound by the roots.
The test compounds are formulated, e.g., by bead-milling with aqueous Dispersol T or as a solution in acetone/ethanol which is diluted to the required concentration immediately before use. 100 p.p.m. a.i. suspensions are sprayed on to the foilage and applied to the roots of the same plant via the soil. (Sprays are applied to maximum retention, and root drenches to a final concentration equivalent to approximately 40 ppm a.i./dry soil). Tween 20, to give a final concentration of 0.1%, is added when the sprays are applied to the cereals.
For most of the tests, the test compound is applied to the soil (roots) and to the foliage (by spraying) one or two days before the plant is inoculated with the diseases. An exception is the test on Erysiphe graminis, in which the plants are inoculated 24 hours before treatment. After inoculation, the plants are put into an appropriate environment to allow infection to take place and then incubated until the disease is ready for assessment. The period between inoculation and assessment typically varies from 4 to 14 days according to the disease and environment.
Chemical Description and Terminology
The compounds of the invention have asymmetric centers; this invention includes all of the optical isomers and mixtures thereof.
Compounds of the invention with carbonxe2x80x94carbon double bonds occur in Z- and E-forms; all isomeric forms of the compounds are included in the invention.
When any variable occurs more than one time in Formula I, its definition on each occurrence is independent of its definition at every other occurrence.
By xe2x80x9cC1-C6 alkylxe2x80x9d or in the invention is meant straight or branched chain alkyl groups or cycloalkyl groups having 1-6 carbon atoms, such as, for example, methyl, ethyl, propyl, isopropyl, n-butyl, sec-butyl, tert-butyl, pentyl, 2-pentyl, isopentyl, neopentyl, hexyl, 2-hexyl, 3-hexyl, and 3-methylpentyl. Preferred C1-C6 alkyl groups are methyl, ethyl, propyl, butyl, cyclopropyl, cyclopropylmethyl, cyclohexyl, cycloheptyl, norbornyl, and the like. Particularly preferred alkyl groups are methyl and ethyl.
By xe2x80x9cC1-C6 alkoxyxe2x80x9d in the invention is meant an alkyl group of indicated number of carbon atoms attached through an oxygen bridge such as, for example, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy, pentoxy, 2-pentyl, isopentoxy, neopentoxy, hexoxy, 2-hexoxy, 3-hexoxy, and 3-methylpentoxy. Preferred alkoxy groups herein are C1-C4 alkoxy groups. Particularly preferred alkoxy groups are ethoxy and methoxy.
The term xe2x80x9chalogenxe2x80x9d includes fluorine, chlorine, bromine, and iodine. Where X is halogen in Formula I-Formula V, bromine is particularly preferred.
xe2x80x9cHaloalkylxe2x80x9d is intended to include both branched and straight-chain saturated aliphatic hydrocarbon groups having the specified number of carbon atoms, substituted with 1 or more halogen atoms. Examples of haloalkyl include, but are not limited to, mono-, di-, or tri-fluoromethyl, mono-, di-, or tri-chloromethyl, mono-, di-, tri-, tetra-, or penta-fluoroethyl, and mono-, di-, tri-, tetra-, or penta-chloroethyl. Typical haloalkyl groups are trifluoromethyl and difluoromethyl. Preferably not more than 5, and more preferably not more than 3 haloalkyl groups, are present in compounds of the invention.
xe2x80x9cHaloalkoxyxe2x80x9d represents a haloalkyl group as defined above with the indicated number of carbon atoms attached through an oxygen bridge.
Non-toxic xe2x80x9cpharmaceutically acceptable saltsxe2x80x9d include, but are not limited to salts with inorganic acids such as hydrochloride, sulfate, phosphate, diphosphate, hydrobromide, and nitrite or salts with an organic acid such as malate, maleate, fumarate, tartrate, succinate, citrate, acetate, lactate, methanesulfonate, p-toluenesulfonate, 2-hydroxyethylsulfonate, salicylate and stearate. Similarly, pharmaceutically acceptable cations include, but are not limited to sodium, potassium, calcium, aluminum, lithium and ammonium. The invention also encompasses the prodrugs of the compounds of Formula I.
Pharmaceutical Preparations
Those skilled in the art will recognize various synthetic methodologies that may be employed to prepare non-toxic pharmaceutically acceptable prodrugs of the compounds encompassed by Formula I. Those skilled in the art will recognize a wide variety of non-toxic pharmaceutically acceptable solvents that may be used to prepare solvates of the compounds of the invention, such as water, ethanol, mineral oil, vegetable oil, and dimethylsulfoxide.
The compounds of general Formula I may be administered orally, topically, parenterally, by inhalation or spray or rectally in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles. Oral administration in the form of a pill, capsule, elixir, syrup, lozenge, troche, or the like is particularly preferred. The term parenteral as used herein includes subcutaneous injections, intradermal, intravascular (e.g., intravenous), intramuscular, spinal, intrathecal injection or like injection or infusion techniques. In addition, there is provided a pharmaceutical formulation comprising a compound of general Formula I and a pharmaceutically acceptable carrier. One or more compounds of general Formula I may be present in association with one or more non-toxic pharmaceutically acceptable carriers and/or diluents and/or adjuvants and if desired other active ingredients. The pharmaceutical compositions containing compounds of general Formula I may be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion, hard or soft capsules, or syrups or elixirs.
Compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents selected from the group consisting of sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients that are suitable for the manufacture of tablets. These excipients may be for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia, and lubricating agents, for example magnesium stearate, stearic acid or talc. The tablets may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monosterate or glyceryl distearate may be employed.
Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example peanut oil, liquid paraffin or olive oil.
Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydropropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide, for example, lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene sorbitan monooleate. The aqueous suspensions may also contain one or more preservatives, for example ethyl, or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose or saccharin.
Oily suspensions may be formulated by suspending the active ingredients in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. The oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide palatable oral preparations. These compositions may be preserved by the addition of an anti-oxidant such as ascorbic acid.
Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already mentioned above. Additional excipients, for example sweetening, flavoring and coloring agents, may also be present.
Pharmaceutical compositions of the invention may also be in the form of oil-in-water emulsions. The oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example liquid paraffin or mixtures of these. Suitable emulsifying agents may be naturally-occurring gums, for example gum acacia or gum tragacanth, naturally-occurring phosphatides, for example soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol, anhydrides, for example sorbitan monoleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monoleate. The emulsions may also contain sweetening and flavoring agents.
Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative and flavoring and coloring agents. The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents that have been mentioned above. The sterile injectable preparation may also be sterile injectable solution or suspension in a non-toxic parentally acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer""s solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.
The compounds of general Formula I may also be administered in the form of suppositories, e.g., for rectal administration of the drug. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient that is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. Such materials are cocoa butter and polyethylene glycols.
Compounds of general Formula I may be administered parenterally in a sterile medium. The drug, depending on the vehicle and concentration used, can either be suspended or dissolved in the vehicle. Advantageously, adjuvants such as local anesthetics, preservatives and buffering agents can be dissolved in the vehicle.
For administration to non-human animals, the composition may also be added to the animal feed or drinking water. It will be convenient to formulate these animal feed and drinking water compositions so that the animal takes in an appropriate quantity of the composition along with its diet. It will also be convenient to present the composition as a premix for addition to the feed or drinking water.
Dosage levels of the order of from about 0.1 mg to about 140 mg per kilogram of body weight per day are useful in the treatment of the above-indicated conditions (about 0.5 mg to about 7 g per human patient per day). The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration. Dosage unit forms will generally contain between from about 1 mg to about 500 mg of an active ingredient.
Frequency of dosage may also vary depending on the compound used and the particular disease treated. However, for treatment of most disorders, a dosage regimen of 4 times daily or less is preferred. For the treatment of eating disorders, including obesity, a dosage regimen of 1 or 2 times daily is particularly preferred. For the treatment of impotence a single dose that rapidly reaches effective concentrations is desirable.
It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, and rate of excretion, drug combination and the severity of the particular disease undergoing therapy.
Preferred compounds of the invention will have desirable pharmacological properties. Such properties include, but are not limited to oral bioavailability, low toxicity, low serum protein binding and desirable in vitro and in vivo half-lifes. Penetration of the blood brain barrier for compounds used to treat CNS disorders is necessary, while low brain levels of compounds used to treat periphereal disorders are often preferred.
Assays may be used to predict these desirable pharmacological properties. Assays used to predict bioavailability include transport across human intestinal cell monolayers, including Caco-2 cell monolayers. Toxicity to cultured hepatocyctes may be used to predict compound toxicity. Penetration of the blood brain barrier of a compound in humans may be predicted from the brain levels of the compound in laboratory animals given the compound intravenously.
Serum protein binding may be predicted from albumin binding assays. Such assays are described in a review by Oravcovxc3xa1, et al. (Journal of Chromatography B (1996) volume 677, pages 1-27).
Compound half-life is inversely proportional to the frequency of dosage of a compound. In vitro half-lifes of compounds may be predicted from assays of microsomal half-life as described by Kuhnz and Gieschen (Drug Metabolism and Disposition, (1998) volume 26, pages 1120-1127).
Compounds of Formula I exhibit good activity in standard in vitro MCH receptor binding assays and/or calcium mobilization assays, specifically in the assays as specified in Examples 5 and 6, which follow. References herein to xe2x80x9cstandard in vitro receptor binding assayxe2x80x9d are intended to refer to that protocol as defined in Example 5 which follows. References herein to xe2x80x9cstandard MCH 1 receptor calcium mobiliztion assayxe2x80x9d are intended to refer to that protocol as defined in Example 6 which follows. Generally, preferred compounds of Formula I have an K1 of about 1 micromolar or less, still more preferably a Ki of about 100 nanomolar or less even more preferably a K1 of about 10 nanomolar or less or even 1 nanomolar or less in such a defined standard in vitro MCH 1 receptor binding assay and exemplied by Example 5. Generally preferred compounds of Formula I are MCH 1 receptor antagonists and exhibit EC50 values of about 4 micromolar or less, more preferably 1 micromolar or less, still more preferably EC50 values of about 100 nanomolar or less even more preferably an EC50 value of about 10 nanomolar or less or even 1 nanomolar or less in such a defined standard in vitro MCH 1 receptor mediated calcium mobilization assay as exemplified by Example 6 which follows.
Preferred compounds of Formula I do not interact with dopamine receptors, particularly human dopamine D2 and D4 receptors. Dopamine receptor binding assays may be preformed using the methods described in Example 9 which follows. Preferred compounds of Formula I exhibit K1 values greater than 1 micromolar in standard assays of dopamine receptor binding assays such as the dopamine D2 and D4 receptor binding assays described in Example 9.